This research project will work towards building a better understanding of the electrochemical behavior of NO and S-nitrosothiols (RSNO's) under biologically relevant conditions. No is believed to act as a biochemical messenger in the regulation of blood pressure and in neurotransmission. In addition, NO is an important cytotoxic agent generated as part of an immune response. RSNO derivatives have been identified as important biochemical "sinks" for NO in serum and cytosol. The results of electrochemical experiments at chemically modified and bare carbon electrodes will be applied in the design and evaluation of microsensors capable of detecting NO and RSNO's in biological matrices such as living tissue and culture media. The development of microsensor capable of analytically monitoring the concentration of these species in high resolution, spatial fashion would be very useful in physiological studies. Previous work by the principal investigator and coworkers has shown that glass carbon (GC) electrodes 3 mm in diameter coated with an iron porphyrin polymer film can be used to detect NO at mild applied potentials. The combination of the mild applied potentials and the iron porphyrin film make this GC/iron porphyrin film electrode highly selective for NO. Oxidizable species such as ascorbic acid, catecholamines, and nitrite did not respond at the applied potential; however, some response to a RSNO derivative was detected. This research project will attempt to miniaturize the iron porphyrin modified electrode using carbon fiber microelectrodes 15 um in tip diameter. Two methods for the attachment of the iron porphyrin films to carbon fiber microelectrodes will be investigated: electrochemical polymerization and direct covalent bonding to the carbon surface. The response of the microsensors to NO and RSNO's will be examined by flow injection analysis. In some additional experiments aimed at better understanding the mechanism of detection, the reaction of water soluble iron-nitrosyl porphyrin adducts with dioxygen will be investigated. The rate of this reaction is believed to be key in maintaining the sensitivity of the GC/iron porphyrin film electrode. Finally, an in-depth electrochemical study of a variety of biologically interesting RSNO derivative and thiols in the presence of NO will be initiated. These experiments will provide some additional insight into the mechanism for RSNO formation in vivo and possibly lead to new ideas for the design of a sensor selective for RSNO derivatives.